EP3082466B1 - Utilisation de thylakoïdes pour réduire l'envie pressante d'aliment appétissant - Google Patents

Utilisation de thylakoïdes pour réduire l'envie pressante d'aliment appétissant Download PDF

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EP3082466B1
EP3082466B1 EP14824825.5A EP14824825A EP3082466B1 EP 3082466 B1 EP3082466 B1 EP 3082466B1 EP 14824825 A EP14824825 A EP 14824825A EP 3082466 B1 EP3082466 B1 EP 3082466B1
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thylakoids
food
isolated
content
hunger
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EP3082466A2 (fr
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Per-Åke ALBERTSSON
Charlotte ERLANSON ALBERTSSON
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THYLABISCO AB
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/66—Microorganisms or materials therefrom
    • A61K35/74—Bacteria
    • A61K35/748—Cyanobacteria, i.e. blue-green bacteria or blue-green algae, e.g. spirulina
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/21—Amaranthaceae (Amaranth family), e.g. pigweed, rockwort or globe amaranth
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
    • A23L33/105—Plant extracts, their artificial duplicates or their derivatives
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/30—Dietetic or nutritional methods, e.g. for losing weight
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
    • A23L33/40—Complete food formulations for specific consumer groups or specific purposes, e.g. infant formula
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/0087—Galenical forms not covered by A61K9/02 - A61K9/7023
    • A61K9/0095—Drinks; Beverages; Syrups; Compositions for reconstitution thereof, e.g. powders or tablets to be dispersed in a glass of water; Veterinary drenches
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/10—Dispersions; Emulsions
    • A61K9/107—Emulsions ; Emulsion preconcentrates; Micelles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/18—Antipsychotics, i.e. neuroleptics; Drugs for mania or schizophrenia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/30—Drugs for disorders of the nervous system for treating abuse or dependence
    • A61P25/32—Alcohol-abuse
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/30—Drugs for disorders of the nervous system for treating abuse or dependence
    • A61P25/34—Tobacco-abuse
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/04—Anorexiants; Antiobesity agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
    • A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs

Definitions

  • the present invention relates to the field of compositions affecting the urge for various types of food, especially palatable food, and the reward value associated with intake of such food.
  • BMI body mass index
  • palatable food e.g. sweet food, or food with high glycemic index, often rich in fat
  • palatable food e.g. sweet food, or food with high glycemic index, often rich in fat
  • Palatable food i.e. sweet and/or fat food, reinforces their intake by increasing the rewarding value and by increasing the pleasure of eating.
  • High-risk eating episodes are those that contain palatable food, like food rich in fat and sugar. To help individuals suffering from overweight and obesity, it is therefore crucial to find ways to strengthen satiety signals and dampen hunger signals.
  • Hedonic hunger (the appetitive drive to eat to obtain pleasure in the absence of an energy deficit), is associated with overeating and with loss of control over eating ( Witt et al International Journal of Eating Disorders Vol. 47(3), p. 273-280, 2014 ). Hedonic hunger favours energy-dense palatable food, rich in sugar and fat, for example snacks, pastries, desserts, baked confectionery and sweets - foods typically ingested in between meals and preferred by women ( Drewnowski, A. (1997). Taste preferences and food intake. Annual review of nutrition, 17, 237-253 )
  • Effective treatment and prevention of obesity should preferably include strategies to suppress the urge for palatable food, especially sweet food. Of interest would further be to dampen the hedonic hunger and the wanting of such food, whereby reducing overeating and assisting in attaining a more healthy eating behavior. Further, also strategies for controlling food-reward behaviors would be of interest, as recognized by Dickson et al (J Neurosci 2012, 32(14):4812-4820 ). However, today no such strategies are available.
  • the present invention seeks to mitigate, alleviate, circumvent or eliminate at least one, such as one or more, of the above-identified deficiencies in the art.
  • Such use of isolated thylakoids will include intake of isolated thylakoids to provide the effect.
  • the intake will provide increased release of GLP-1 (glucagon-like peptide-1), whereby the rewarding effect of palatable food, affecting the reward center in the brain, is decreased.
  • GLP-1 glycol-like peptide-1
  • the present invention is based on the use of thylakoids in man, wherein the use includes intake of isolated thylakoids.
  • Thylakoids are membrane-bound compartments present inside chloroplasts of plants and inside cyanobacteria.
  • the thylakoid is the site of the light-dependent reactions of photosynthesis.
  • the light-independent reactions of photosynthesis i.e. the Calvin-Benson Cycle
  • carbon dioxide is converted to three-carbon sugars, which later are combined to form sucrose and starch. This process is often denoted carbon fixation.
  • Thylakoids consist of a thylakoid membrane surrounding a thylakoid lumen. In the chloroplast, thylakoids are commonly grouped as stacks of disks being refereed to as grana.
  • Thylakoids may be extracted from the chloroplast of green leaves and from cyanobacteria. Their content includes proteins, lipids and pigments (e.g. chlorophyll and carotenoids).
  • the general principles of isolating and enriching thylakoids have been described elsewhere (cf. PCT/SE2006/000676 , WO/ SE2009/000327 , and Emek, SC, Szilagyi A, Akerlund H E, et al. (2010) A large scale method for preparation of plant thylakoids for use in body weight regulation Prepar biochem & biotech 40 (1), 13-27 ).
  • the cells of green leaves are broken mechanically.
  • the thylakoids are then separated from the cell walls and other large cell fragments by filtration.
  • Thylakoids are then collected from the filtrate by precipitation at pH 4.7 and washed in water by repeated centrifugations.
  • Thylakoids have previously been found to increase satiety, decrease homeostatic hunger signals and promote weight loss in several studies, both in human and animal subjects (cf. WO 2006/132586 and WO 2010/008333 among others). Thylakoids, or parts thereof, may be included in foodstuff in many different ways since they are themselves food. They may also be included in different types of pharmaceutical formula as previously described in WO 2006/7132586 .
  • green leaves naturally comprise thylakoids
  • intake of fresh green leaves does not provide the same effect as intake of isolated thylakoids, or parts thereof.
  • Plant cells comprising thylakoids are surrounded by very thick and tough cell wall. Although we chew a lot when eating vegetables, not many cells are broken. In order to dissolve the cell walls, special enzymes such as cellulase and pectinas etc. are required. However, these enzymes are not present in the stomach or in the small intestine of humans. Unless extracted from the greens leaves, the thylakoids may not exert any effect in the small intestine before reaching the large intestine, wherein the cell walls may be broken down by bacteria.
  • an important aspect of obesity is the tendency of obese individuals not only to overeat but also to preferentially choose palatable food, e.g. food with high glycemic index (i.e. 60 or above) often with high fat content, as well as sweet tasting food comprising sweeteners or sugars, and sweet and fat food, over healthy food.
  • palatable food e.g. food with high glycemic index (i.e. 60 or above) often with high fat content, as well as sweet tasting food comprising sweeteners or sugars, and sweet and fat food, over healthy food.
  • Fast-food e.g. pizza, hamburgers, French fries, soft drinks etc.
  • the preferential for palatable food is linked to the rewarding value of the food; over-eating may thus be driven by the wish to obtain pleasure.
  • reducing the urge for palatable food is extremely important in order for a weight reduction program to be successful.
  • not only obese subject would have use of a composition providing reduced urge for palatable food, but also normal weight subjects.
  • Reducing the hedonic hunger may facilitate in attaining a more healthy eating behavior.
  • decreasing the rewarding effect of palatable food may also facilitate in attaining a more healthy eating behavior.
  • Attenuating the hedonic drive in those who experience increased cravings for palatable food is a way of avoiding overeating and weight gaining.
  • Intake of food having high glycemic index or tasting sweet, as well as food having a high fat, content is associated with increased risk for obesity, cardiovascular diseases, including coronary diseases, and diabetes, as demonstrated in several studies (cf. Johnson RK, Appel LJ, Brands M, Howard BV, Lefevre M, Lustig RH, Sacks F, Steffen LM, Wylie-Rosett J: Dietary sugars intake and cardiovascular health: a scientific statement from the American Heart Association.
  • An important factor for the preference for palatable food is the reward value linked to it. Eating palatable food results in release of reward factors having an important role in over-eating. could the effect of these reward factors be reduced, the preference for palatable food and also over eating would be reduced.
  • the present inventors have surprisingly revealed that intake of isolated thylakoids not only affects the lipolytic activity and thereby the appetite (homeostatic hunger - Fig. 1A ), but also, and importantly, reduces the urge for palatable food (hedonic hunger - Figs. 1B-D among others). Interestingly, the finding is not related to the suppression of appetite, as the effect is being present also in hungry individuals (cf. Fig. 1A vs. Figs. 1B-D ). In other words, the reduced urge for palatable food is observed not only immediately after eating but also in the late postprandial phase for at least six hours when hunger is present.
  • GLP-1 glucagon-like peptide-1
  • GLP-1 not only affects the reward value of food but also of other stimulants including alcohol
  • PLoS One 2013, 8(4):e61965 It is believed that the mechanism is related to suppression of dopamine in the reward centers of the brain.
  • thylakoids In contrast to the previous uses of thylakoids, relying on local effects exerted by thylakoids in the intestines, the present use is believed to relate to central effects exerted by thylakoids affecting the reward center of the brain. Further, the effect is also distinct from the increased levels of CCK//decreased levels of ghrelin previously reported.
  • An embodiment of the present invention therefore relates to the use of isolated thylakoids to suppress, lower, decrease, reduce, lessen, and/or ease the urge for palatable food, in order to decrease hedonic hunger, in man.
  • Such use may be non-therapeutic. Further, such use may be therapeutic.
  • the use includes intake of isolated thylakoids. While the person taking isolated thylakoids will be less prone to select palatable food over more healthy alternatives, the person will not all the suddenly dislike palatable food.
  • the reduced urge for palatable food will assist in treating and preventing of obesity, as persons taking isolated thylakoids will be less prone to overeat. Further, the reduced urge for palatable food may assist in maintaining the weight after successfully having taken part in a weight loss program.
  • palatable food examples include sweets, pastries, e.g. doughnuts, croissants, shortbread cookies and wafers, candy, potato chips, French fries, soft drinks, e.g. Coca Cola, Fanta, Sprite, Pepsi Cola, and energy drinks (e.g. Red Bull), chocolate, e.g. milk chocolate and chocolate bars, fast food, e.g. pizza and hamburgers, and ice-cream.
  • sweets pastries, e.g. doughnuts, croissants, shortbread cookies and wafers, candy, potato chips, French fries, soft drinks, e.g. Coca Cola, Fanta, Sprite, Pepsi Cola, and energy drinks (e.g. Red Bull), chocolate, e.g. milk chocolate and chocolate bars, fast food, e.g. pizza and hamburgers, and ice-cream.
  • sweets pastries
  • croissants e.g. doughnuts, croissants, shortbread cookies and wafers
  • candy e.g.
  • potato chips
  • palatable food has a high glycemic index and/or tastes sweet.
  • sweet food not having a high GI
  • e.g. food comprising fructose or a sweetener may trigger a reward effect and hence be involved in over eating.
  • soft drinks also provides a rewarding effect, which indirectly contributes to over-eating. Diet drinks do cause a release of dopamine, demonstrating that they also have a rewarding effect that can lead to overeating.
  • food having a moderate GI of at least 50 and high fat content such as French fries, will trigger a reward effect and may hence be involved in over eating.
  • salty snacks with high fat content, such as salted nuts trigger a reward effect.
  • sugars present in the palatable food in amounts herein specified above are selected from the group consisting of the monosaccharides glucose (also known as dextrose), fructose and galactose and the disaccharides sucrose, maltose and lactose, and mixtures thereof.
  • Another embodiment relates to the use of isolated thylakoids, or parts thereof, to decrease the rewarding effect of food, such as palatable food.
  • Such use may be non-therapeutic. Further, such use may be therapeutic. Examples of palatable food have been provided herein above.
  • a method of suppressing, lowering, decreasing, reducing, lessening, and/or easing the urge for palatable food comprises the step of administering isolated thylakoids, or parts thereof, to a subject, e.g. a human being, whose urge for palatable food is to be affected.
  • thylakoids As already described, intake of thylakoids, or parts thereof, has also been shown to reduce the need or urge to smoke in man; probably by affecting the reward center of the brain. Not only palatable food and nicotine is affecting the reward center of the brain, but also other stimulants, such as alcohol, i.e. ethanol.
  • a further embodiment relates to the use of isolated thylakoids, or parts thereof, to release GLP-1 (glucagon-like peptide-1) in man and to a method of releasing GLP-1 (glucagon-like peptide-1) in man including administration of isolated thylakoids, or parts thereof.
  • Such use may be non-therapeutic. Further, may such use be therapeutic.
  • GLP-1 is known to decrease the rewarding effects of various stimuli, including palatable food, nicotine and alcohol in the reward center in the brain.
  • Isolated thylakoids, or parts thereof may thus also be used to decrease the rewarding effect of stimulants affecting the reward center in the brain.
  • isolated thylakoids, or parts thereof may be used in a method of decreasing the rewarding effect of stimulants affecting the reward center in the brain.
  • Thylakoids may be isolated from plants as well as from cyanobacteria. Typically, the thylakoids are enriched from green leaves or green algae. The thylakoids may origin from leaves of any photosynthesizing plants, such as clover, rape, sugar beet, dandelion, Arabidopsis thaliana, maize, tobacco, sun flower, Chenopodium, Atriplex, spinach, mangold, quinoa, kale, sugar beet and grasses. Preferably, the thylakoids are isolated and enriched from spinach. The isolated thylakoids among other parts of the thylakoid, comprises thylakoid membranes, or parts thereof. Methods for isolating thylakoids have been discussed herein above.
  • the thylakoids may be isolated and concentrated in a manner such that the chlorophyll content of the isolated thylakoids is from about 8 mg to 150 mg, such as 10-100 mg, chlorophyll per g isolated thylakoids.
  • the isolated thylakoids have a chlorophyll a/b ratio of from about 2.0 to 4.0, preferably about 2.6 to 3.4, and most preferably about 2.9 to 3.4.
  • the protein:chlorophyll ratio in the isolated thylakoids may be from about 3.0 to about 10.0, preferably about 2.0 to 5.0, 3.0 to 5.0, or 6.0 to 8.0.
  • the thylakoids are administered orally to provide their effect. While they may be administered directly, they typically are administered as an integral part of a composition, such as nutraceutical composition, e.g. a foodstuff or a dietary supplement, or pharmaceutical composition, e.g. a medicament.
  • a composition such as nutraceutical composition, e.g. a foodstuff or a dietary supplement, or pharmaceutical composition, e.g. a medicament.
  • the medicament except for the isolated thylakoids, also comprises at least one pharmaceutical acceptable excipient, such as a carrier, a diluent, and/or a stabilizer.
  • pharmaceutically acceptable relates to an excipient that, at the dosage and concentration employed, does not cause any unwanted effects in the subject to whom it is administered.
  • pharmaceutically acceptable excipients are well-known in the art.
  • a medicament as disclosed herein may also comprise other pharmaceutically acceptable excipients, such as preservatives, antioxidants, colouring agents and the like.
  • the isolated thylakoids are added as food additive to a foodstuff, to provide a food stuff having the ability to affect the reward center in the brain, as described herein. Similar to the isolated thylakoids such food stuff finds none-therapeutic use.
  • the isolated thylakoids are added as an ingredient to a dietary supplement, to provide a dietary supplement having the ability to affect the reward center in the brain, as described herein. Similar to the isolated thylakoids, such a dietary supplement may find therapeutic as well as none-therapeutic use.
  • the dietary supplement may for example be a fruit juice shoot, e.g. a blueberry shot, or a vegetable soup or shoot, e.g. potato soup, broccoli soup, potato soup, or tomato soup, in which the isolated thylakoids, or parts thereof, are added.
  • the ingredients in the dietary supplement are such that contain antioxidants, vitamins, fibers which give further usefulness as to health.
  • the content of thylakoids in the composition may correspond to a chlorophyll content of 1 mg to 100 mg chlorophyll per g composition (dry weight).
  • the composition comprises from about 8 to about 80 mg chlorophyll per gram (dry weight), preferably from about 10 to about 50 mg, such as about 10 mg to 30 mg, 20 mg to 40 mg, or 30 mg to 40 mg chlorophyll content per gram (dry weight).
  • the carbohydrate content of the composition may be from about 5 to about 50 g carbohydrate content per 100 g of the composition (dry weight), preferably about 10 g to 40 g per 100 g, such as 15 g to 30 g, 20 g to 30 g, or 30 g to 40 g per 100 g (dry weight).
  • the protein content of the composition may be from about 10 to about 60 g protein per 100 g of the composition (dry weight), preferably about 10 g to 40 g per 100 g, preferably 15 g to 30 g, 20 g to 30 g, or 30 g to 40 g per 100 g (dry weight).
  • the composition is an oral pharmaceutical or nutraceutical composition comprising a physiologically tolerable oil-in-water emulsion comprising thylakoids.
  • the amount of thylakoids may be 20 to 30 wt.%, such as about 25 wt.%.
  • the oil-content may be 20 to 30 wt.%, such as about 25 wt.%.
  • the oil used may be vegetable oils of food grade, such as edible rape seed oil (also known as Canola oil), olive oil, sunflower oil etc.
  • the daily dose of thylakoids to be administered may vary.
  • the amount of the composition to be administered may be selected in manner such that the dosage correlates to a daily dose of 2.4 mg to 240 mg thylakoids per kg body mass of the mammal.
  • a composition as disclosed herein therefore further comprises probiotic bacteria.
  • the probiotic bacteria to be included in the composition may be Bifidobacterium, e.g. Bifidobacterium infantis, or Lactobacillus, e.g. Lactobacillus plantarum.
  • prebiotic agents such as trans-galactooligosaccharide, inulin, fructooligosaccharide (FOS), lactulose and mannan Oligosaccharides (MOS), may be included in the composition.
  • Example 1 decreased urge for palatable food
  • FFM Fat Free Mass; includes bones, muscles and organs.
  • a 50 g blueberry shot with or without supplementation of 5 g thylakoids were taken approximately five minutes before their breakfast. The subjects were told to eat all given food within 15 minutes. No drinking or eating was permitted between meals.
  • a second meal consisting of a pizza (Grandiosa Gudfadern, Procordia AB, Eslöv, Sweden), water and coffee/tea were given to subjects after 6 hours with the instruction to eat and drink until satisfied. The second meal was served in a cubicle for each subject one by one.
  • Blood samples were taken just before breakfast (time point: 0 minute) and thereafter at 15, 30, 45, 60, 90, 120, 180, 240, 300 and at 360 minutes. The lunch was given just after the blood sampling at time point 360 minute. Analysis was carried out for a number of components including GLP-1.
  • VAS Visual Analogue Scale
  • the questionnaire was also answered between the second and third meal at time points 60, 120, 180 and 240 minutes after lunch.
  • the thylakoid powder used in the present meal study was produced as Appethyl, by Future-ceuticals, Chicago, IL, USA. 5 gram of the powder were mixed with 50 g blueberry juice (Ekströms Bl ⁇ bärssoppa Orginal, Procordia AB, Eslöv, Sweden) and 2 g rapeseed oil (Zeta rapsolja, DILUCA & DI LUCA AB, Sweden). The control was consisting of 50 g blueberry juice and 2 g rapeseed oil.
  • the plasma GLP-1 analys was made with an enzyme linked immunosorbent assay (ELISA) kit (Cat. # EGLP-35K; Millipore Corp. Billerica, MA, USA).
  • the kit analyses only the biologically active forms of GLP-1: GLP-1 (7-36 amide) and GLP1 (7-37 amide) in human plasma.
  • a fluorescence plate reader FLUOstar OPTIMA 413-101; BMG LABTECH GmbH, Ortenberg, Germany
  • the inter-assay coefficient of variation (CV) is between 1-13 % and intra-assay between 6-9 %, accordning to the manufactures data.
  • VAS visual Analogue Scale
  • the diet regime i.e. intake of 5 g thylakoids per day
  • the diet regime continued for three months.
  • thylakoids have the ability to suppress urge or desire for sweet, salt and fat. This suppression of desire lasts the whole day following consumption of a thylakoid shot in the morning. Furthermore, the liking of palatable food decreased further after 3 months of thylakoid treatment.
  • the effect on hunger was much less pronounced (cf. Fig. 1A ).
  • the women had significantly less desire for palatable food, such as candy, chocolate or potato chips.
  • the effect underlying the present invention is thus apparently not related to the previous findings that intake of enriched, isolated thylakoids may suppress appetite, due to decreased lipolytic activity (cf. WO 2006/132586 ), and delay uptake of molecules, such as glucose (cf. WO 2012/113918 ).
  • glucagon-like peptide GLP-1
  • the thylakoids thus cause endogenous release of a hormone that regulates reward-related eating behavior in a direction that is helpful to prevent and treat obesity.
  • subjects having taken thylakoids will be more prompted to choose a healthy meal as the urge for palatable food is significantly reduced. Further, the risk for over-eating is reduced. This will lead to a decreased risk for type-2-diabetes and cardiovascular disease associated with obesity and insulin resistance.
  • Example 2 decreased need for nicotine (not within the scope of the invention)
  • thylakoid shoots are provided as part of a weight loss program.
  • a man typically smoking 15 to 20 cigarettes a day, reported that his consumption of cigarettes was reduced to less than 10 cigarettes per day - typically to about 9 cigarettes per day - 10 days after he had started taking thylakoid shoots regularly in the morning.
  • Example 3 decreased urge for palatable food during dietary intervention
  • example 1 The effects observed in example 1 were achieved without any caloric restriction. However, most weight loss programs are based on a caloric restriction. Hence, the effect of thylakoids on hedonic hunger during a caloric restriction commonly used in weight loss studies was also studied.
  • Baseline characteristics of the 26 included subjects are listed in Table 1. One participant in the control group was not included in the analysis of subjective ratings of hunger, satiety and urges for specific food items due to non-attendance. Table 1. Baseline characteristics of the 26 women included in the study (average ⁇ SD). There were no significant differences between the groups.
  • the participants were randomized into two groups to ensure normal distribution within and between the groups based on body weight, BMI, blood glucose, insulin, triacylglycerides (TAG) and cholesterol (total and LDL).
  • One of the groups received supplementation by thylakoids in a blueberry drink every day, while the other group served as control, receiving a daily blueberry drink without thylakoids.
  • an isocaloric breakfast (2114 kJ/505 kcal) consisting of the blueberry drink with or without thylakoids, yoghurt with apple, breakfast cereal, nuts and coffee or tea was served ( Table 2 ).
  • an isocaloric take-away lunch 2593 kJ/630 kcal consisting of a frozen thai-curry meal, bean salad and a banana was administered.
  • VAS questionnaires measuring subjective parameters of hunger, satiety and urge for specific foods were filled out at given time points throughout the day on both day 1 and 56, whereas blood samples were taken only during the initial 240 minutes following breakfast. Table 2. Contents of breakfast and lunch served on first and last days of the study. Meal components Amount (g) Calories (kJ/kcal) Protein (g) Fat (g) Carbohydrates (g) Fiber (g) Breakfast Natural yoghurt 300 753 / 180 10.2 9.0 14.4 0.0 Muesli wt.
  • the thylakoids used in the present study were prepared from baby spinach leaves using the pH-method, as described ( Emek SC, et al A large scale method for preparation of plant thylakoids for use in body weight regulation, Prep Biochem Biotechnol 2010, 40(1):13-27 ).
  • the thylakoid-slurry was dried to obtain a thylakoid powder, prepared by Swepharm AB (Södra Sandby, Sweden).
  • 100g thylakoids consists of 41.1 g protein, 14.5 g fat, 36.8 g carbohydrate, 3.5 g salt as well as pigments such as 3640 mg chlorophyll, 28 mg lutein, 730 ug zeaxantin, 4 760 ug betakaroten, 21 ug vitamin A, 1330 ug vitamin K, 6.07 mg vitamin E and 166 ug folic acid.
  • the thylakoid group received 5.6 g of thylakoid powder mixed with 2.8 g rapeseed oil (Zeta, Di Luca & Di Luca AB, Sweden) and 50 g of blueberry soup (Ekströms original, Procordia Food AB, Eslöv, Sweden).
  • the control group received 2.8 g rapeseed oil mixed with 50 g blueberry soup.
  • the blueberry drinks with and without thylakoids contained 209 kJ/50 kcal versus 188 kJ/45 kcal respectively. The drinks were taken before breakfast every day.
  • Subjects were also instructed to accomplish 60 minutes of low/medium intensity exercise each day, such as power walking, swimming, basic aerobics etc.
  • Body weight was measured with a digital scale (TANITA WB-100A, class III, Amsterdam, The Netherlands).
  • the composition of body fat mass (kg) and trunk fat (kg) were measured with a Bioelectric Impedance Analyser (TANITA-BC 418 MA). Waist circumference, midway between the lower rib margin and the iliac crest, and hip circumference were measured to the nearest 0.5 cm by using a non-stretchable tape measure.
  • VAS Ant A, et al Reproducibility, power and validity of visual analogue scales in assessment of appetite sensations in single test meal studies, Int J Obes Relat Metab Disord 2000, 24(1):38-48 .
  • VAS Flint A, et al Reproducibility, power and validity of visual analogue scales in assessment of appetite sensations in single test meal studies, Int J Obes Relat Metab Disord 2000, 24(1):38-48 .
  • Pictures assisted the evaluation of the urge for specific food items.
  • For high carbohydrate snack pictures of a sandwich and a sweet cinnamon bun were presented, for salt and fat; pictures of potato chips and salted peanuts, for sweet snack; pictures of candy and a popsicle and for fat and sweet; pictures of cake and chocolate were presented.
  • thylakoids has an effect on hedonic hunger, being distinct from its long term effect on homeostatic hunger, confirming the effect of the present invention. Further, as is apparent from example 1 and 3, thylakoids are able to suppress hedonic hunger, irrespective of caloric restriction.
  • intake of thylakoids alleviates some of the strains coupled to caloric restriction during body weight loss programs. Further, intake of thylakoids assists in adopting a more healthy eating behavior and reducing the relative intake of palatable food. This may be crucial in maintaining a healthy eating behavior and avoiding weight gain subsequent to a weight loss program.
  • isolated thylakoids are able to suppress hunger hedonic hunger irrespective of caloric restriction.
  • TANITA-BC 418 MA Amsterdam, The Netherlands.
  • the study was a randomised, placebo-controlled, double blind, single-centred meal intervention study with a cross over design, conducted at the Biomedical Centre (BMC) at Lund University, Sweden. Each participant was tested on two days, separated by a wash out period of at least 1 week, receiving a green-plant supplement (5 g) on one day and placebo on the other. The allocation to treatment or placebo was randomised and balanced between test days.
  • VAS Visual Analogue Scales
  • the VAS-questionnaire was filled out every hour as well as before and after lunch and the ad libitum snack buffet, served at 13:00 and 16:00 respectively. Between breakfast and lunch, participants left the premises for work or free activities, and were reminded to fill out the VAS-questionnaires every hour via text messages. After lunch, the participants stayed in their allocated seats, sitting side by side with cardboard dividers between them, all facing a window. They were allowed to read or work on laptops until 16:00 when the snack buffet was served at the table on individual trays. A leaflet, containing information about the products served, accompanied the buffet, ensuring the freshness of the foods. It included names of manufacturers and pictures of the packaging. The same pictures were used for the VAS-questionnaire.
  • the participants Before starting to eat from the snack buffet in front of them, the participants filled out the VAS-questionnaire once more. Then they were allowed to eat freely from the products on their trays, encouraged to follow their urges and ask for more should they run out. They were occupied by the snack buffet for 45 min, not doing anything else during this time. Afterwards, they filled out the VAS-questionnaire again, this time including questions regarding how much they liked the products they tasted. Finally, they had the opportunity to comment freely on how they felt during the test day.
  • the number of participants required to achieve a power of 0.8 in this crossover study was calculated using data from an earlier study. Based on these calculations, 15 subjects were needed to detect a 10 mm difference in VAS-ratings of hunger. This number of subjects is also sufficient for detecting differences in ratings for wanting palatable food. According to literature, 17 subjects should be sufficient for detecting a difference in food intake of 500kJ/120kcal.
  • the green-plant membranes (Appethyl, Greenleaf Medical AB, Sweden) used in the present study were prepared from baby spinach leaves using the pH-method as previously described ( Emek et al., Prep Biochem Biotechnol, 40, 13-27 ), followed by drum drying.
  • 100 g of green-plant membranes contain 26.1 g protein, 7.24 g fat, 48.7 g carbohydrate, 27.9 mg lutein, 730 ⁇ g zeaxantin, 3.45 mg betakaroten, 21 ⁇ g vitamin A, 1330 ⁇ g vitamin K 1 , 6.07 mg vitamin E and 166 ⁇ g folic acid.
  • Breakfast on both test days was a standardized Swedish high carbohydrate breakfast with yoghurt and muesli, one slice of white bread with butter, cheese and sweet pepper, coffee or tea.
  • the energy distribution was 31.2 per cent of total energy (E%) fat, 15.4 E% protein and 53.4 E% carbohydrates. Sugars constituted 22.6 E%.
  • the lunch served on both test days was a slice of pizza, made from sourdough, with tomato sauce, ham, onion, mushrooms and cheese, served with boiled broccoli and tap water. Coffee or tea following lunch was optional, but subjects were required to have the same both test days. Energy distribution of the lunch was 45.9 E% carbohydrates, 32.5 E% fat and 21.6 E% protein.
  • a range of bestselling snacks, sweets and confectionery was chosen for the snack buffet (Table 3), based on the responses to the product questionnaire. These products were grouped in three different categories: salty, sweet, and sweet-and-fat snacks. Snacks with salty taste included potato chips and salted assorted nuts, products with sweet taste, containing sugar, included two different kinds of sweets and orange juice, products with sweet taste containing both sugar and fat were: milk chocolate, dark chocolate, chocolate covered toffee, chocolate pastry, and cinnamon bun. Each participant received a tray with the products served in separate containers, weighed before and after consumption. The amounts were chosen to be more than enough for one person, to ensure ad libitum consumption. Coffee, tea and water were served with the snacks in a limited amount.
  • the drinks were optional, but the subjects had to have the same on both test days.
  • Table 3 Components of the snack buffet, energy content per 100g Products Calories (kJ/kcal) Protein (g/E%) Fat (g/E%) Carbohyd r.
  • the product questionnaire was constructed for this study as a tool for choosing well-suited products to measure cravings and snack intake in a Swedish population. It contained specific questions about snacks of the most popular brands in Sweden: potato chips, nuts, chocolate, sweets, baked confectionery, buns, fruit juice and soft drinks. Questions were asked about how often the responder consumed the palatable foods, how much they liked the products on a nine point hedonic scale and which product in each category they preferred.
  • the TFEQ-R18V2 is a revised version of the original Three-Factor Eating Questionnaire (TFEQ), a self-assessment scale widely used in studies of eating behaviour. While the original TFEQ contained 51 items, TFEQ-R18V2 contains 18 questions to measure three types of eating behaviour: emotional eating, uncontrolled eating and cognitive restraint. The questionnaire used in the present study was a Swedish translation.
  • TFEQ Three-Factor Eating Questionnaire
  • VAS visual analogue scales
  • the questionnaire was constructed as a booklet with written instructions on the first page. All questions were followed by a 100-mm horizontal line, on which subjects marked their response. The line was anchored at each end, expressing the minimum value on the left: “Not at all”, and the maximum value on the right: “Extremely”. At the last time point of the day, questions were added about liking for the products tasted that particular day, to enable evaluation of the rewarding properties of the items.
  • ratings for the individual food items were merged into three different categories and presented as salty, sweet, and sweet-and-fat snacks (Table 3). For analyses regarding all snacks together, all values were merged and presented as total/all snacks.
  • Treatment effect of green-plant membranes on feelings of hunger, satiety and urge for specific food products and liking respectively was calculated by subtracting VAS-ratings for treatment day from control day values, except for satiety, which was calculated the opposite way.
  • Treatment effect on food intake was calculated as difference in caloric intake between control and treatment days.
  • Calculating delta values for the treatment effects was required to account for paired observations. These delta values were then correlated to eating behaviour scores, as measured by the TFEQR18-V2 questionnaire, divided into the three domains: emotional eating, uncontrolled eating and cognitive restraint. Correlations were also calculated between eating behaviour scores, BMI and body fat percentage. All correlations were computed by Pearson correlation coefficient.
  • Wanting for sweet-and-fat decreased 36% in the treated group compared to control ( p ⁇ 0.05, tAUC, Fig. 7F ).
  • the urge for sweet-and-fat snacks was lower in the treated group compared to control at time points 60, 120, 180 minutes and also right before lunch at 285 minutes ( Fig. 7E ).
  • the treatment effect on wanting sweet-and-fat snacks as well as all kinds of snacks showed positive correlation with emotional eating scores, so that higher scores for emotional eating behaviour was correlated to a greater treatment effect of green-plant membranes on ratings for wanting palatable food in general, and sweet-and-fat foods in particular (p ⁇ 0.01 respectively, Pearson r, Fig. 10A and B ).
  • Cognitive restraint and uncontrolled eating domains failed to show any correlation between treatment effects and score for eating behaviour.
  • the present example demonstrates that intake of thylakoids prior to breakfast increase subjective ratings of satiety and decrease ratings of hunger as well as cravings for snacks and sweets during the day.
  • intake of thylakoids prior to breakfast increase subjective ratings of satiety and decrease ratings of hunger as well as cravings for snacks and sweets during the day.
  • eating behaviour scores treatment effect of green-plant membranes on wanting was positively correlated to emotional eating for all snacks, sweet-and-fat foods being specifically targeted.
  • Intake of thylakoids also decreased subjective liking for sweet, scored directly following consumption.
  • Treatment also had an attenuating effect on hunger ratings throughout the day. In comparison to ratings for satiety, the difference in hunger ratings reached significance later, at 180 minutes after breakfast in comparison to 60 minutes for satiety.
  • the present findings on hunger verify previous findings on decreased hunger ratings following treatment with green-plant membranes.
  • the suppressed hunger may be related to reduced ghrelin-levels and an indirect effect of increased circulating levels of the satiety hormones CCK and GLP-1.
  • the later appearance of hunger suppression suggests an indirect effect by thylakoids on promotion of satiety hormones.
  • the suppressed hedonic hunger by thylakoids may be an effect of altered levels of the reward-related appetite hormone GLP-1.
  • Findings on reduced liking are important, since liking of sweetness influence wanting of sweet-tasting products. Indeed, overweight women have been shown to have an increased liking for sweet. A reduced liking may, over time, cause a reduction in wanting for sweet. As a consequence the reduced wanting may decrease snacking in between meals and thus overeating.
  • treatment reduced wanting for sweet to a greater extent compared to other kinds of snacks. There was also an earlier onset. Wanting for sweet was reduced from 15 minutes and onwards, in comparison to wanting for salty and sweet-and-fat snacks, which were reduced from 60 minutes respectively. This implies that acute treatment with green-plant membranes in particular curbes the sweet taste. The mechanism for this targeting is not known.
  • the present example shows that treatment with green-plant membranes attenuate hunger, homeostatic and hedonic, as well as wanting for palatable food and liking for sweet. Reducing wanting is important, since wanting is a major cause of hedonic eating, which contributes to overconsumption and obesity. In addition, individuals who are obese and/or dieting are even more susceptible to hedonic hunger. Therefore, reducing cravings for palatable food is necessary, both to control appetite, prevent weight gain and to facilitate a permanent weight loss. Supplementation by green-plant membranes may help prevent eating between meals, hence over-eating and in the long run, overweight.

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Claims (13)

  1. Utilisation non thérapeutique de thylacoïdes isolés pour réduire l'envie pressante d'aliment appétissant, de manière à réduire la faim hédonique, chez l'homme, dans laquelle ladite utilisation inclut la prise de thylacoïdes isolés.
  2. Utilisation selon la revendication 1, dans laquelle l'aliment appétissant présente une ou plusieurs des caractéristiques suivantes :
    - un indice glycémique d'au moins 60, par exemple d'au moins 70 ;
    - une teneur en sucre élevée d'au moins 5 % en poids pour les boissons et d'au moins 20 % en poids, par exemple d'au moins 50 % en poids, pour un autre aliment ;
    - comprend un édulcorant ;
    - une teneur en graisse élevée d'au moins 20 % en poids et une teneur en sucre élevée d'au moins 20 % en poids ;
    - une teneur en graisse élevée d'au moins 20 % en poids et un indice glycémique d'au moins 50 ; et
    - une teneur en graisse élevée d'au moins 20 % en poids et une teneur en sel (NaCl) d'au moins 1,0 % en poids.
  3. Utilisation selon la revendication 2, dans laquelle l'aliment appétissant présente au moins l'une des caractéristiques suivantes :
    - une teneur en sucre élevée d'au moins 5 % en poids pour les boissons et d'au moins 20 % en poids, par exemple d'au moins 50 % en poids, pour d'autres aliments ; et
    - une teneur en graisse élevée d'au moins 20 % et une teneur en sucre élevée d'au moins 20 % en poids ;
    dans laquelle les sucres compris dans l'aliment appétissant sont sélectionnés dans le groupe constitué des monosaccharides glucose, fructose et galactose, des disaccharides sucrose, maltose et lactose, et de mélange de ceux-ci.
  4. Utilisation selon l'une quelconque des revendications 1 à 3, dans laquelle la libération du GLP-1 (peptide de type glucagon 1) est accrue par ladite utilisation de thylacoïdes isolés.
  5. Utilisation selon l'une quelconque des revendications 1 à 4, dans laquelle l'effet de satisfaction de l'aliment appétissant, affectant le centre de satisfaction dans le cerveau, est réduit par ladite utilisation de thylacoïdes isolés.
  6. Utilisation non thérapeutique de thylacoïdes isolés pour réduire l'effet de satisfaction d'un aliment appétissant, par exemple d'un aliment appétissant selon l'une quelconque des revendications 2 ou 3.
  7. Utilisation selon l'une quelconque des revendications 1 à 6, dans laquelle les thylacoïdes ont été isolés de feuilles vertes de plantes ou d'algues vertes ; de préférence les thylacoïdes ont été isolés de feuilles de trèfle, navette, betterave à sucre, pissenlit, Arabidopsis thaliana, maïs, tabac, tournesol, Chenopodium, Atriplex, épinard, betterave champêtre, quinoa, chou frisé ou d'herbes ; de manière préférée entre tous les thylacoïdes ont été isolés de l'épinard.
  8. Utilisation selon l'une quelconque des revendications 1 à 7, dans laquelle les thylacoïdes isolés font partie intégrante d'une composition nutraceutique.
  9. Utilisation selon la revendication 8, dans laquelle la teneur en thylacoïdes isolés dans la composition nutraceutique correspond à une teneur en chlorophylle d'environ 10 à environ 50 mg de chlorophylle par gramme de la composition (poids sec) ;
    de préférence la teneur en glucides de la composition nutraceutique est de 5 à 50 g de glucides pour 100 g de la composition nutraceutique (poids sec) ; et/ou
    la teneur en protéine de la composition nutraceutique est de 10 à 60 g de protéine pour 100 g de la composition nutraceutique (poids sec).
  10. Utilisation selon la revendication 8 ou 9, dans laquelle la composition nutraceutique comprend une émulsion huile-dans-l'eau physiologiquement tolérable comprenant des thylacoïdes isolés.
  11. Utilisation selon la revendication 10, dans laquelle la composition nutraceutique comprend des bactéries probiotiques et facultativement un agent prébiotique.
  12. Utilisation selon la revendication 5, dans laquelle l'effet de satisfaction est réduit par augmentation de la libération de GLP-1 (peptide de type glucagon 1).
  13. Utilisation selon la revendication 1, dans laquelle la faim hédonique est réduite par augmentation de la libération de GLP-1 (peptide de type glucagon 1).
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